PO.ET02.08 · 实验与分子治疗
首个基于凝聚层的递送系统用于干细胞中高效且安全的基因工程
The first coacervate based delivery system for efficient and safe genetic engineering in stem cells
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
干细胞生物学协调胚胎发生、影响衰老，并为革命性疗法带来希望，是人类健康的核心主线。这凸显了对先进递送技术的迫切需求，以促进研究进展和临床转化。在此，我们利用凝聚层——在进化中被视为原细胞——开发了一个开创性的递送平台。我们鉴定出一种哺乳动物内源性蛋白，它通过液-液相分离（LLPS）包裹多种核酸，形成基于蛋白质的凝聚层。这些凝聚层表现出优异的mRNA包封能力、高效的细胞内化、强大的胞质载荷释放，以及跨多种细胞类型（包括原代免疫细胞和干细胞）的广泛兼容性。由此产生的平台命名为ProteanFect，可支持多种遗传载荷以实现多功能的基因操作策略。ProteanFect在关键干细胞类型中展现出高度的多功能性和高效性，包括造血干细胞（HSCs）、人胚胎干细胞（ESCs）以及来源于多种体细胞来源（如PBMCs、尿液和脂肪细胞）的诱导多能干细胞（iPSCs）。它通过递送mRNA或Cas9 RNP复合物高效地实现了基因过表达和敲除。作为其治疗血液系统疾病潜力的关键展示，ProteanFect在HSCs中的BCL11A位点实现了约80%的等位基因修饰，这是一种旨在重新激活胎儿血红蛋白以治疗血红蛋白病的策略，且未检测到脱靶编辑。此外，该平台通过将先导编辑工具递送至PBMC来源的iPSCs中促进了精确的基因校正，实现了约55%的精确碱基替换并保持高细胞活力。总之，ProteanFect构成了一个多功能、安全且高效的非病毒平台，无需合成脂质或电穿孔即可运作。通过在不发生基因组整合的情况下实现重编程和基因编辑载荷的强大瞬时表达，它为细胞重编程、iPSC生成以及下一代细胞疗法的开发提供了一条更安全、更可控的途径。
查看英文原文 English abstract
Stem cell biology, which orchestrates embryogenesis, influences aging, and holds promise for revolutionary therapies, represents a central thread in human health. This underscores the pressing need for advanced delivery technologies to facilitate both research progress and clinical translation. Here, we harness coacervates-regarded as protocells during evolution-to develop a pioneering delivery platform. We identified a mammalian endogenous protein that encapsulates diverse nucleic acids through liquid-liquid phase separation (LLPS), forming protein-based coacervates. These coacervates exhibit superior mRNA encapsulation capacity, efficient cellular internalization, robust cytoplasmic cargo release, and broad compatibility across multiple cell types -including primary immune cells and stem cells. The resulting platform, named ProteanFect, supports diverse genetic cargoes for versatile gene manipulation strategies. ProteanFect demonstrated high versatility and efficiency across key stem cell types, including hematopoietic stem cells (HSCs), human embryonic stem cells (ESCs), and induced pluripotent stem cells (iPSCs) derived from diverse somatic sources such as PBMCs, urine, and adipocytes. It efficiently enabled gene overexpression and knockout through the delivery of mRNA or Cas9 RNP complexes. In a key demonstration of its therapeutic potential for hematological disorders, ProteanFect achieved approximately 80% allele modification at the BCL11A locus in HSCs, a strategy aimed at reactivating fetal hemoglobin to treat hemoglobinopathies, with no detectable off-target editing. Furthermore, the platform facilitated precise genetic correction by delivering prime editing tools into PBMC-derived iPSCs, achieving approximately 55% precise base substitution with high cell viability. In summary, ProteanFect constitutes a versatile, safe, and efficient non-viral platform that operates without synthetic lipids or electroporation. By enabling robust transient expression of reprogramming and gene-editing cargo without genomic integration, it provides a safer and more controllable pathway for cellular reprogramming, iPSC generation, and the development of next-generation cell therapies.
利益披露 Disclosure
P. Zhu, None.
M. Lu,
Nanoportal Biotech Employment.
Q. Zhang, None..
R. Zhang, None.
L. Jiang,
Nanoportal Biotech Employment.
X. Fei,
Nanoportal Biotech Employment.
X. Gao,
Nanoportal Biotech g., Board of Directors, non-salaried role).